Ultrasensitive and Highly Selective Detection of <i>Staphylococcus aureus</i> at the Single-Cell Level Using Bacteria-Imprinted Polymer and Vancomycin-Conjugated MnO<sub>2</sub> Nanozyme
Yixin Ma, Xiaohui Lin, Bin Xue, Donglei Luan, Chunping Jia, Shilun Feng, Xiaojun Bian, Jianlong Zhao
Abstract
Pathogenic bacterial infections, even at extremely low concentrations, pose significant threats to human health. However, the challenge persists in achieving high-sensitivity bacterial detection, particularly in complex samples. Herein, we present a novel sandwich-type electrochemical sensor utilizing bacteria-imprinted polymer (BIP) coupled with vancomycin-conjugated MnO 2 nanozyme (Van@BSA-MnO 2 ) for the ultrasensitive detection of pathogenic bacteria, exemplified by Staphylococcus aureus ( S. aureus ). The BIP, in situ prepared on the electrode surface, acts as a highly specific capture probe by replicating the surface features of S. aureus . Vancomycin (Van), known for its affinity to bacterial cell walls, is conjugated with a Bovine serum albumin (BSA)-templated MnO 2 nanozyme through EDC/NHS chemistry. The resulting Van@BSA-MnO 2 complex, serving as a detection probe, provides an efficient catalytic platform for signal amplification. Upon binding with the captured S. aureus, the Van@BSA-MnO 2 complex catalyzes a substrate reaction, generating a current signal proportional to the target bacterial concentration. The sensor displays remarkable sensitivity, capable of detecting a single bacterial cell in a phosphate buffer solution. Even in complex milk matrices, it maintains outstanding performance, identifying S. aureus at concentrations as low as 10 CFU mL –1 without requiring intricate sample pretreatment. Moreover, the sensor demonstrates excellent selectivity, particularly in distinguishing target S. aureus from interfering bacteria of the same genus at concentrations 100-fold higher. This innovative method, employing entirely synthetic materials, provides a versatile and low-cost detection platform for Gram-positive bacteria. In comparison to existing nanozyme-based bacterial sensors with biological recognition materials, our assay offers distinct advantages, including enhanced sensitivity, ease of preparation, and cost-effectiveness, thereby holding significant promise for applications in food safety and environmental monitoring.